目的构建精氨酸甲基转移酶2(PRMT2)慢病毒表达载体。方法通过PCR扩增PRMT2 c DNA,将PRMT2 c DNA连接于GV308载体,经测序确认后,将GV308/PRMT2与p Helper 1.0和p Helper 2.0共转染至293T细胞中,收获病毒,通过Real time PCR测定滴度;将PR...目的构建精氨酸甲基转移酶2(PRMT2)慢病毒表达载体。方法通过PCR扩增PRMT2 c DNA,将PRMT2 c DNA连接于GV308载体,经测序确认后,将GV308/PRMT2与p Helper 1.0和p Helper 2.0共转染至293T细胞中,收获病毒,通过Real time PCR测定滴度;将PRMT2慢病毒表达载体侵染293T细胞,通过四环素诱导和Western blot检测PRMT2慢病毒表达载体的表达能力。结果在感染PRMT2慢病毒载体293T细胞中能检测到PRMT2-3Flag融合蛋白的表达。结论成功构建PRMT2的慢病毒表达载体。展开更多
Histone arginine methylation by protein arginine methyltransferases(PRMTs)is crucial for transcriptional regulation and is implicated in cancers.Despite their therapeutic potential,some PRMTs present challenges as dru...Histone arginine methylation by protein arginine methyltransferases(PRMTs)is crucial for transcriptional regulation and is implicated in cancers.Despite their therapeutic potential,some PRMTs present challenges as drug targets due to their context-dependent activities.Here,we demonstrate that hypoxia triggers the rapid condensation of PRMT2,which is essential for its histone H3R8 asymmetric dimethylation(H3R8me2a)activity.This process depends on PRMT2's integration into transcriptional condensates,which is mediated by phosphorylation at Serine 12 within its N-terminal intrinsically disordered region.This phosphorylation is critical for hypoxia-inducible gene expression and glioblastoma(GBM)progression.Transcription-associated cyclin-dependent kinases(CDKs),particularly CDK9,drive PRMT2S12 phosphorylation.Inhibition of CDK9 using TG02 suppresses hypoxia-induced H3R8me2a and transcriptional activity.Moreover,the combination of TG02 and temozolomide,the standard chemotherapy for GBM,significantly inhibits tumor progression in mouse xenograft models,an effect partially mediated by targeting PRMT2S12 phosphorylation.Our study uncovers the role of transcriptional condensation in enhancing PRMT activity,reveals a new mechanism for CDK9 inhibitors in modulating context-dependent transcriptional programs,and proposed a combinatorial therapeutic strategy against GBM.展开更多
文摘目的构建精氨酸甲基转移酶2(PRMT2)慢病毒表达载体。方法通过PCR扩增PRMT2 c DNA,将PRMT2 c DNA连接于GV308载体,经测序确认后,将GV308/PRMT2与p Helper 1.0和p Helper 2.0共转染至293T细胞中,收获病毒,通过Real time PCR测定滴度;将PRMT2慢病毒表达载体侵染293T细胞,通过四环素诱导和Western blot检测PRMT2慢病毒表达载体的表达能力。结果在感染PRMT2慢病毒载体293T细胞中能检测到PRMT2-3Flag融合蛋白的表达。结论成功构建PRMT2的慢病毒表达载体。
基金supported by the National Natural Science Foundation of China(32320103009,82473964)Postdoctoral Fellowship Program of China Postdoctoral Science Foundation(GZC20231923,2024M752388,2025T180713)+1 种基金Tianjin Municipal Science and Technology Project(24JCQNJC00980)the National Youth Talent Support Program to X.W.
文摘Histone arginine methylation by protein arginine methyltransferases(PRMTs)is crucial for transcriptional regulation and is implicated in cancers.Despite their therapeutic potential,some PRMTs present challenges as drug targets due to their context-dependent activities.Here,we demonstrate that hypoxia triggers the rapid condensation of PRMT2,which is essential for its histone H3R8 asymmetric dimethylation(H3R8me2a)activity.This process depends on PRMT2's integration into transcriptional condensates,which is mediated by phosphorylation at Serine 12 within its N-terminal intrinsically disordered region.This phosphorylation is critical for hypoxia-inducible gene expression and glioblastoma(GBM)progression.Transcription-associated cyclin-dependent kinases(CDKs),particularly CDK9,drive PRMT2S12 phosphorylation.Inhibition of CDK9 using TG02 suppresses hypoxia-induced H3R8me2a and transcriptional activity.Moreover,the combination of TG02 and temozolomide,the standard chemotherapy for GBM,significantly inhibits tumor progression in mouse xenograft models,an effect partially mediated by targeting PRMT2S12 phosphorylation.Our study uncovers the role of transcriptional condensation in enhancing PRMT activity,reveals a new mechanism for CDK9 inhibitors in modulating context-dependent transcriptional programs,and proposed a combinatorial therapeutic strategy against GBM.
